Care and Use of Nalgene Labware

For most general applications, Thermo Scientific™ Nalgene™ plastics may be washed in a mild (non-alkaline is recommended) detergent, followed by a rinse with tap water and then distilled water. Do not use abrasive cleaners or scouring pads on any plastic labware. Do not use strong alkaline cleaning agents with polycarbonate.

As the chart below shows, for most trace metal analyses, plastic is generally "cleaner" or less contaminated than glass or other materials. However, plastic does contain trace levels of certain metals. To minimize potential low-level contamination, remove these metals or leach them from plastic by soaking in 1N HCl and rinsing in distilled water. For extremely precise work, use HCl, followed by soaking in 1N HNO3, and rinsing in distilled water. Soaking time may vary according to individual needs, but plastic should be soaked no longer than 8 hours. If more rigorous cleaning is desired, increase the concentration of acids used. Caution: concentrated nitric acid is a strong oxidizing agent and will embrittle many plastics.

To remove trace organics which contribute to trace metal absorption, clean plastic surfaces with alcohol, alkalies, alcoholic alkalies or chloroform. A final rinse of 1N HCl also minimizes absorption of trace elements.

Polysulfone (PSF), a resin used in Thermo Scientific™ Nalgene™ centrifuge tubes and reusable filterware, is extremely "cleanable." The following qualification testing was performed on Nalgene PSF centrifuge tubes at The University of Rhode Island's Graduate School of Oceanography (URI-GSO).

URI-GSO was determining the concentration of trace metals in atmospheric aerosols and seawater from remote marine locations. They measured the input to the Pacific Ocean of heavy metals and other trace elements; pesticide, petroleum and plastic residues; other natural and man-made organic materials, as well as compounds causing acid rain. The trace metal concentrations in the samples were extremely low, so any labware used needed to be extremely cleanable to prevent leaching of contaminants from the vessels or filtration equipment into the samples. In addition, the labware could not irreversibly adsorb metals from sample solutions. The labware in question included filter holders, which were used during procedures for extracting various trace metals from the seawater samples.

This cleaning procedure was generated from the procedures used at URI-GSO, J.R. Moody and R.M. Lindstrom Analytical Chemistry 49:2264 (1977) and conversations with faculty of the California Institute of Technology.

The following testing procedure was used:

Approximately 25 mL 1N quartz redistilled HNO3 was put into five of the centrifuge tubes.

Approximately 25 mL of 1 part per billion aluminum, copper, lead and zinc in 1 N quartz redistilled HNO3 was put into the other five centrifuge tubes.

The tubes were allowed to stand for one month, and the contents of the tubes were analyzed.

The analyses were performed by atomic absorption spectrophotometry on a Perkin Elmer 5000 equipped with an HGA 5000 utilizing Zeeman background correction. All standards used are traceable back to NBS (National Bureau of Standards).

The results and conclusions were:

(1) The 1N quartz redistilled HNO3 had immeasurably low concentrations of metals (e.g., aluminum, copper and lead 0.1 parts per billion; zinc 0.01 parts per billion). Therefore, once cleaned, polysulfone leaches insignificant amounts of these four metals into solution.

(2) The 1 part per billion aluminum, copper, lead and zinc solutions had, within experimental error, 1 part per billion aluminum, copper, lead and zinc. Therefore, polysulfone does not adsorb these metals from an acidified solution.

The analysis of aluminum, copper, lead and zinc involved a two-step extraction procedure. The first, a "liquid/liquid" extraction with an organic complexing agent, removed copper, lead and zinc. The second employed an iron hydroxide co-precipitation technique. Iron nitrate was added to the seawater and the pH was adjusted. Iron hydroxide and aluminum were filtered from the rest of the mixture. This is where the Nalgene reusable filter holders, molded of PSF, were used.

Summary of Average Element Content of 12 Plastics and Borosilicate Glass

Greases and Oils - General Cleaning

We recommend using non-alkaline detergents for cleaning plastic labware, especially those products made of polycarbonate, which is particularly sensitive to alkaline attack.

Nalgene L900 Liquid Detergent (Cat. No. 900-4000) is designed to clean all plastics and glassware at a neutral pH. A 5% solution in water is usually sufficient but can be increased to 20% for stubborn residue or heavily soiled labware. L900 Detergent can be used in automatic washers for lightly to normally soiled items.

Soak the labware in the detergent for up to 3 hours, then gently wash with a cloth or sponge. Soak heavily soiled items in a 5 to 20% concentration in water for 4 or more hours prior to washing. Rinse with tap water and then distilled water.

Do not use abrasive cleaners or scouring pads on any plastic labware.

Periodically disassemble and clean spigots and threads on bottles and closures to prevent salt build-up, which can cause leakage.

Most plastics, particularly the polyolefins (LDPE, HDPE, PP, PMP, and PPCO) have non-wetting surfaces that resist attack and are easy to clean.

For many applications, washing with a mild detergent will remove greases and oils. When more rigorous cleaning is needed, organic solvents may be used with caution. Extended exposure to these solvents may cause some swelling of polyolefins. Rinse off all solvents before using labware. Use only alcohols on PC, PSF, PS, and PVC; other organic solvents will attack these plastics. Do not use organic solvents with acrylic.

Dishwashers

Labware washing machines can be used with all resins except LDPE, acrylic and PS, due to temperature limitations.

Special note on polycarbonate (PC) Repeated washings in the dishwasher weaken the exceptional strength of PC. PC labware that has been exposed to high stresses (such as those caused by centrifugation or use in vacuum chambers) should always be washed by hand using a mild, neutral pH, non-abrasive detergent without sheeting agents, such as Nalgene L-900.

Keep the dishwasher cycle time to a minimum. Use the plastics cycle and set the water temperature at 135°F (57°C) or lower. Remove the labware as soon as possible after cooling is complete. Avoid excessive abrasion of plastics by covering metal spindles with soft material such as plastic tubing. Plastic labware should be weighted down and held in place with accessory racks.

Ultrasonic Cleaners

Ultrasonic cleaning units may be used to clean labware as long as the labware does not rest directly on the transducer diaphragm.

Organic Matter

Chromic acid solution will remove organic matter, but will eventually embrittle plastics. To minimize embrittlement, soak plastic for no more than 4 hours. The following formula is the recommended cleaning agent:

This solution is designed to produce an excess of dichromate in the form of a precipitate which actually extends the useful life of chromic acid and dissolves as needed. This chromic acid solution can be used repeatedly until it begins to develop a greenish color, indicating a loss of potency. As a result of the excess dichromate built into this formula, the solution lasts much longer than commercially available solutions.

Sodium hypochlorite solutions (bleach) are also effective in removing organic matter. Use at room temperature.

How To Remove RNase or DNase From Plastic Containers

RNase, an enzyme that breaks down RNA, and DNase, which breaks down DNA, are contaminants that can interfere with nucleotide research. DNase can be destroyed by autoclaving for 15 minutes at 121°C or by following any of the procedures listed below. One or more of the following techniques will inhibit or remove RNase from your plastic container. Match the resin code on the bottom of your Nalgene container with the correct technique.

Heat at 180°C for at least 8 hours.1

Rinse in chloroform.1

Soak in a 0.1% aqueous solution of diethyl pyrocarbonate2 (DEPC) for 2 hours at 37°C; rinse several times with sterile (DEPC-treated) water(***); heat to 100°C for 15 minutes OR autoclave for 15 minutes at 121°C on a liquid/slow exhaust cycle. (Heating or autoclaving will remove DEPC residues.) Note heating variations in the following chart.

Clean equipment with a detergent solution, rinse thoroughly with water and rinse with 95% ethanol to dry. Soak the equipment in a 3% hydrogen peroxide (H2O2) for 10 minutes at room temperature. Rinse the equipment thoroughly with DEPC-treated water.***

Soak equipment in 0.1N Sodium Hydroxide (NaOH) in 0.1% EDTA in water overnight and then rinse thoroughly with DEPC-treated water.3

Nalgene L900 Liquid Detergent

Thermo Scientific™ Nalgene™ L900 Liquid Detergent is recommended for hand and automatic machine washing of glass and plastic labware use in pharmaceutical, medical, research, academic, biotechnology, environmental, and other laboratories. It is especially friendly to polycarbonate and will not cause clouding, stress cracking, or crazing like alkaline detergents.

Specification

Inhibition Residue Test Results show no toxic or inhibiting effects caused by residue from L900 liquid detergent (differences in averaged counts on the plates were less than 15% as outlined in Standard Methods, 18th edition); no alkaline or acidic residue left by L900 detergent, indicating no pH effect. pH is 6 to 7.

Wash Instructions

Hand wash use:

For lightly soiled items use 2–4 mL per liter of water. For heavily soiled items use up to 15 mL per liter of water allow to soak for 4 or more hours before washing.

Automatic washer use:

Follow the washer manufacturer's directions for liquid detergent use. When in doubt, use 2–4 mL per liter of water. Heavily soiled items may need to be presoaked. The amount of detergent needed may vary from washer to washer, depending on the amount of water used in each wash cycle.

Centrifuge Ware Use and Cleaning

Inspection Guidelines

Inspect centrifuge tubes and bottles carefully before each use.

Centrifuge ware is subjected to high g-forces while spinning, which can lead to failure. Safe laboratory practice requires that all centrifuge ware be inspected before each use. Plastic centrifuge ware is easy to inspect; it requires no special equipment. Initially, the effect of excessive stress in plastic can be seen as cloudiness and discoloration or as "crazing", i.e. minute cracks visible when the tube is held at an angle in front of a bright light. With continued use, a crazed tube will develop larger cracks or will fail. Tubes should be discarded if cracks are readily visible to the unaided eye.

Rotor Balancing

Consult your centrifuge operator's manual for instructions on rotor balancing and handling. Proper rotor handling, cleaning, and balancing are extremely important. Because plastics have different densities, tubes and bottles of different styles or materials should not be arranged randomly in the rotor. All centrifuge ware must be at least 80% full for proper performance, unless otherwise noted.

Cleaning

Disposable Tubes

Disposable centrifuge ware should be discarded after one use. NOTE: When handling hazardous materials, decontaminate tubes prior to disposal. To clean reusable Nalgene centrifuge ware, we suggest the following procedure:

To loosen any residuals, presoak the tubes or bottles in Nalgene L900 detergent (see General Cleaning for specific recommendations). Soak overnight to loosen stubborn residue.

Remove residue with a non-abrasive brush, or with a rubber or fluoropolymer policeman.

Wash and rinse product thoroughly, with distilled water as a final rinse.

Air dry.

Sterilization

PP, PPCO, PMP, FEP and ETFE products can be autoclaved repeatedly under normal conditions, 15 minute cycle at 121°C/15 psig (1.02 bar). PC and PSF products can be autoclaved under these same conditions, but autoclaving will cause deterioration in mechanical strength and will shorten their usable life. If you autoclave PC or PSF products, follow the Inspection Guidelines above. When autoclaving Nalgene centrifuge ware, follow these guidelines:

Clean and rinse tubes or bottles thoroughly with distilled water before autoclaving.

For bottles or tubes with closures, remove closures (cap and plug) completely before autoclaving to prevent collapse of container when cooling.

Certain chemicals which have no appreciable effect on resins at room temperatures may cause deterioration at autoclaving temperatures. See the Centrifuge Ware Chemical Resistance Chart for specific chemicals and resins. All Nalgene centrifuge ware can also be sterilized with ethylene oxide gas or compatible chemical disinfectants.

Effects of Chemicals

Chemicals can affect the strength, flexibility, surface appearance, color, dimensions, and weight of plastic parts. Chemical resistance is influenced by temperature, duration, and frequency of exposure, chemical concentration, and centrifugal force. Physical changes which may be caused by chemical exposure include:

Absorption of solvents, resulting in softening or swelling of the plastic

Permeation of solvent through the plastic

Dissolution of polymer in the solvent

Stress-cracking, which may occur as a result of chemical exposure combined with external stress of centrifugal forces on tubes or bottles, which can be worsened by improper fit in a rotor cavity

Nalgene centrifuge tubes and bottles should not break or crack if used in a properly fitting rotor and run according to our recommendations regarding chemical resistance, temperature limits, relative centrifugal force ratings, use of closures, washing, and autoclaving. If the limits are exceeded, the tube or bottle may fail during centrifugation (i.e., may crack or break). However, unlike glass, a Nalgene tube or bottle will not shatter, thus minimizing the risk to users and equipment.

Autoclaving

Recommended autoclave cycle is 121°C, 15 psig (1 bar) for 20 minutes.

All items should be carefully cleaned before autoclaving to prevent baking contaminants onto the surface of the plastic. After cleaning, all items should be rinsed thoroughly in distilled water before autoclaving. Certain chemicals which have no appreciable effect on plastics at room temperature may cause deterioration at autoclaving temperatures and therefore must be removed.

Do not autoclave containers (except those made of fluoropolymers) containing detergent or wetting solutions.

Before autoclaving, just set cap or closure on top of the container without engaging the threads. (If this is not done, pressure differentials may cause containers to collapse during autoclaving.)

For best results, use a slow exhaust cycle.

Carboys with spigots must always be autoclaved empty (the spigot may leak during the autoclave cycle).

Cycle Times

Plastics transfer heat more slowly than glass or metal and may take longer to reach sterilizing temperatures in the autoclave. Because of differences in heat transfer characteristics between plastics and inorganic materials, the contents of plastic containers may take longer to reach sterilization temperature (121°C). Therefore, longer autoclaving cycles are necessary for liquids in large-volume plastic containers. Adequate cycles can be determined only by experience with specific liquids and containers.

Chemical additives in steam will attack transparent plastics and cause a permanently glazed surface after autoclaving.

Some transparent plastics may absorb minute amounts of water vapor and appear cloudy after autoclaving. The clouding will disappear as the plastic dries. Clearing may be accelerated in a drying oven at 110°C.

Use polypropylene copolymer (PPCO) bottles instead of polysulfone with Tween in the autoclave.

Test tube racks filled with tubes must be autoclaved on a flat surface.

Specific Plastic Considerations

Polycarbonate products are autoclavable. They must be thoroughly rinsed before autoclaving because detergent residues cause crazing and spotting. Autoclaving cycles should be limited to 20 minutes at 121°C. PC shows some loss of mechanical strength after repeated autoclaving and therefore may not function well under high-stress applications, such as centrifugation. Our PC vacuum chambers are considered "not autoclavable" for this reason.

Caution: Do not use strong alkaline detergents on polycarbonate. Do not use boiler steam containing alkaline chemical additives that may attack the plastic and cause the item to fail.

Teflon and Tefzel are registered trademarks of DuPont* used under license by Nalge Nunc* International Corporation*.

Acetal products are autoclavable at recommended settings. Proper ventilation is required as acetal will emit formaldehyde odor during autoclaving. The following statement complies with the California Safe Drinking Water and Toxic Enforcement Act of 1986:"WARNING: Upon autoclaving, this product may release formaldehyde, a chemical known to the State of California as a carcinogen."

Polysulfone products are autoclavable. They are somewhat weakened by repeated autoclaving, although less than polycarbonate. If autoclaved repeatedly, polysulfone products will eventually fail under high-stress applications, such as high-speed centrifugation.

Nalgene PVC Tubing can be autoclaved, but ethylene oxide or chemical disinfectant is preferred. If you autoclave it, follow these guidelines:

Clean and rinse tubing thoroughly, including final rinse with distilled or deionized water.

Place on a nonmetallic tray in the autoclave so wrapped tubing is not touching wall or rack of autoclave.

Do not stack anything on the tubing. Use 15 minute cycle at 121°C, 15 psig.

Restore clarity of tubing by drying approximately 2 hours at a temperature no higher than 75°C.

Nalgene Silicone Tubing can be autoclaved for 30 minutes at 121°C, 15 psig in muslin cloth or sterilizing paper.

Labware made of the following plastics is not autoclavable under any conditions: polystyrene, polyvinyl chloride (except PVC tubing), styrene acrylonitrile, acrylic, low-density and high-density polyethylene and polyurethane.

Other Sterilizing Methods

Gas

Polypropylene, polymethylpentane, polypropylene copolymer, Teflon ETFE, Tefzel FEP, PFA, polycarbonate, acetal and polysulfone products as well as PVC, and silicone tubing may be gas-sterilized (ethylene oxide, formaldehyde). We recommend allowing an appropriate aeration time suited to the particular application before re-using the item.

Teflon and Tefzel are registered trademarks of DuPont used under license by Nalge Nunc International Corporation.

Chemical Disinfectants

In general, all of the aforementioned plastics can be subjected to commonly used disinfectants (quaternary ammonium compounds, iodophors, formalin, benzalkonium chloride, ethanol, etc.). There may be some surface attack (crazing) when using a more chemically aggressive disinfectant on the less resistant plastics (PS, SAN, PVC, PC, acrylic) with prolonged use.

Dry Heat

Dry heat sterilization is recommended only for ACL (acetal or polyoxymethylene), Halar ECTFE (ethylene-chlorotrifluoroethylene copolymer), EFTE (ethylene-tetrafluoroethylene), Teflon FEP (fluorinated ethylene propylene), PPCO (polypropylene copolymer), PC (polycarbonate), Teflon PFA (perfluoroalkoxy), PMP (polymethylpentene), PP (polypropylene), and Teflon TFE (tetrafluoroethylene). Maximum temperatures and minimum sterilization times at each temperature for each of these resins are given in the following table:

Teflon is a registered trademark of DuPont used under license by Nalge Nunc International Corporation.

Dry Heat Sterilization

Resins

Temperature

Time

FEP, PFA, PMP*, TFE

170°C (338°F)

60 min.

PSF

160°C (320°F)

120 min.

ECTFE, ETFE

150°C (302°F)

150 min.

140°C (284°F)

180 min.

ACL, PPCO*, PC, PP*

121°C (250°F)

Overnight

* with no load, no stacking

Microwaving

In general, all plastics allow transmission of microwaves. Among commonly used plastics, PMP and PSF are most transparent to microwaves. We recommend their use when the chemical and temperature resistance of PMP and PSF are compatible with the material to be heated. Use fluoropolymers when aggressive chemicals, such as acids or solvents, are being heated; proper venting is essential. Closure threads must be completely disengaged before heating bottles or containers in a microwave oven.

Pre-Sterilized Nalgene Labware

How is the sterility of Nalgene Labware assured?

Some Nalgene labware is sterilized and tested after assembly, and sterility is guaranteed as long as the packaging is intact. Two methods of sterilization are used—ethylene oxide gas and gamma irradiation. Although our disposable filtration products are not classified as medical devices, they are sterilized following Good Manufacturing Practices of the United States FDA Medical Device Act of 1976. American Association of Medical Instrumentation (AMMI) recommended practices are used as a guideline for our sterilization methods.

Indicator dots used on Nalgene products undergo an irreversible color change upon sterilization and are specific for either ethylene oxide gas (EtO) or gamma irradiation. Purple EtO dots turn green when products have been properly processed. Yellow gamma dots turn red when products have been properly exposed.

Ethylene Oxide

Ethylene oxide (EtO) is used for sterilization where low temperature and pressure are necessary (the material to be sterilized cannot be autoclaved). The labware is exposed to EtO in chambers where temperature, humidity, and pressure are carefully controlled and monitored. We use three methods to assess the outcome of the EtO procedure. External color indicators on the packaging demonstrate exposure to the gas but not sterility. Biological indicators are used to evaluate sterility. These indicators are resistant strains of bacterial spores, usually Bacillus subtillis var. niger, whose failure to grow demonstrates the effectiveness of the EtO cycle. Sterility testing may be performed on filterware after the sterilization process. As with biological indicators, the criterion for confirming sterility is the absence of microbial growth. After EtO sterilization, the labware is quarantined until all test results are known. This quarantine period lasts from 7 to 14 days and assures adequate time for the outgassing of EtO and possible residues. Nalgene labware is routinely evaluated for the presence of these residues, and the levels found fall well below the FDA recommended limits for implanted medical devices. Nalgene labware sterilized by EtO include: sterilization filter units, analytical filter units, and filter funnels.

Gamma Irradiation

During this procedure, the labware is exposed to high energy, ionizing gamma radiation from a Cobalt 60 source at room temperature. This process is indicated where there must be an absolute assurance that no potentially toxic residue, however minute, may be present to affect the experimental procedure. Product sterility is achieved by an accumulated absorbed radiation dosage measured in megarads (Mrads). Dosage levels are selected by a process verification dose experiment, which includes bioburden determination and sterility testing. The sterility of the product is assured by dosimetric release, which confirms that the specified minimum dose has been delivered to the product. Nalgene labware sterilized by gamma irradiation includes all of our tissue culture filter units, our filter unit receivers and PETG bottles and carboys.

Other Testing

All Nalgene sterile labware products undergo testing for:

Bioburden

Pyrogens

Cytotoxicity1

Nalgene filtration products are also subjected to several other tests, regardless of the type of sterilization they undergo. These include:

Membrane retention/unit integrity

Membrane thickness

Membrane extractables

Bubble point

Details on these tests are available from Thermo Fisher Scientific Technical Service. Certification of sterility and compliance with specifications will be supplied upon written request. Write to Quality Assurance Department, Thermo Fisher Scientific, Box 20365, Rochester, New York 14602-0365 USA. Or email us at technical.nalgene@thermofisher.com.

Special Cleaning Procedures

For most trace metal analyses, plastic is generally "cleaner" or less contaminated than glass or other materials. However, plastic does contain trace levels of certain metals. To minimize potential low-level contamination, remove these metals or leach them from plastic by soaking in 1N HCl and rinsing in distilled water. For extremely precise work, use HCl, followed by soaking in 1N HNO3, and rinsing in distilled water. Soaking time may vary according to individual needs, but plastic should be soaked no longer than 8 hours. If more rigorous cleaning is desired, increase the concentration of acids used. Caution: concentrated nitric acid is a strong oxidizing agent and will embrittle many plastics.

To remove trace organics which contribute to trace metal absorption, clean plastic surfaces with alcohol, alkalies, alcoholic alkalies or chloroform. A final rinse of 1N HCl also minimizes absorption of trace elements.

Polysulfone (PSF), a resin used in Nalgene centrifuge tubes and reusable filterware, is extremely "cleanable." The following qualification testing was performed on Nalgene PSF centrifuge tubes at The University of Rhode Island's Graduate School of Oceanography (URI-GSO).

URI-GSO was determining the concentration of trace metals in atmospheric aerosols and seawater from remote marine locations. They were measuring the input to the Pacific Ocean of heavy metals and other trace elements; pesticide, petroleum and plastic residues; other natural and man-made organic materials, as well as compounds causing acid rain. The trace metal concentrations in the samples were extremely low, so any labware used needed to be extremely cleanable to prevent leaching of contaminants from the vessels or filtration equipment into the samples. In addition, the labware could not irreversibly adsorb metals from sample solutions. The labware in question included filter holders, which were used during procedures for extracting various trace metals from the seawater samples.

Before labware contaminated with infectious or toxic materials is removed from the work area, it should be sterilized appropriately. Autoclaving is the preferred method for sterilization; however, any method of chemical disinfecting or heat sterilization appropriate for the particular plastic may be used (see Sterilizing Plastics). Liquid waste containing biohazardous materials must always be decontaminated before disposal.

Labware that is contaminated with both biohazardous and radioactive material must first be sterilized. Methods for removing radioactive material depend on the isotope used, its quantity, half-life, material and solubility. For routine decontamination of non-infectious/non-toxic materials, first soak in decontaminant/cleaner for 24 hours at room temperature. Follow with several rinsings in distilled water. To accelerate decontamination, increase the cleaner concentration and solution temperature. Agitation and careful scrubbing with non-abrasive materials will also speed this process. Be particularly careful not to scratch PC. Always dispose of radioactive wastes and effluents properly.